M. tuberculosis infection remains a severe global health crisis that demands new therapeutic agents and preventative strategies. In light of these requirements, the discovery of biological processes required for the virulence of this organism have been paramount for the identification of potential drug targets. A large number of these proteins have been implicated in cell envelope biogenesis and maintenance. Previous work in our laboratory has shown that the Rip1 (Rv2869c) intramembrane protease is required for growth and persistence in mice and is necessary for maintenance of the cell envelope of mycobacteria. Rip1 is a member of the Site two protease (S2P) family of intramembrane metalloproteases which cleave substrates within their transmembrane domains. These proteolytic cascades have been implicated in the transduction of environmental stimuli between cellular compartments via downstream transcriptional networks. Preliminary data show that Rip1 protease activity is activated by iron limitation and controls multiple downstream pathways. This application describes a logical approach in which a combination of conventional genetics and biochemical techniques allow for the identification of the relative contribution of the downstream transcriptional targets to Rip1 dependent attenuation in the mouse. Furthermore, we will determine the molecular mechanism responsible for Rip1 dependent low iron signaling.